Characterization of 1-Aminocyclopropane-1-carboxylate synthase purified from mung bean hypocotyls

De Sheng Tsai, Richard N. Arteca, Jeannette M. Arteca, Allen T. Phillips

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

Analysis of the properties of the key enzyme for ethylene production, 1-aminocyclopropane-1-carboxylate (ACC) synthase, isolated in pure form from hormonally induced etiolated mung bean hypocotyl segments, has revealed that the optimum pH for catalytic activity under normal assay conditions was pH 8.0 although maximal stability was noted at pH 6 to 7. The purified enzyme has an isoelectric point of 5.1 when analyzed by isoelectric focusing and the enzyme's content of pyridoxal phosphate was determined by two separate procedures to be one per subunit of molecular weight 65,000. Edman degradation provided the sequence of 26 residues from the N-terminal portion of the protein: V-A-H-A-K-D-D-A-Y-L-Q-A-A-I-P-K-R-I-K-L-F-E-T-I-Q-A-. Routine assays conducted for 5 minutes with the hydrogen sulfate salt of S-adenosyl methionine (AdoMet) revealed no apparent inhibition. Longer incubation time with substrate, however, indicated that AdoMet was irreversibly inactivating the enzyme and suggested the possibility that this might have important consequences for enzyme turnover. Aminooxyacetic acid was a potent competitive inhibitor of enzyme activity with a Ki of 2 µM. No inhibition was noted with L-homoserine, methylthioadenosine, 5'-AMP, 5'-ADP, 5'-ATP or fusicoccin. An investigation of the methods required to stabilize ACC synthase from mung beans, tomato and apple found that high concentration (>0.5M) of potassium phosphate buffer were effective. Activity remained unchanged during 48-hour storage at 4 °C, and after 10 days in storage, 50 % of the original activity remained for enzyme extracted from mung bean and tomato, while activity from apple tissue was lost after 7 days. Such findings should facilitate the comparative analyses of these important ACC synhases when all have been purified.

Original languageEnglish (US)
Pages (from-to)301-306
Number of pages6
JournalJournal of Plant Physiology
Volume137
Issue number3
DOIs
StatePublished - Jan 1 1991

All Science Journal Classification (ASJC) codes

  • Physiology
  • Agronomy and Crop Science
  • Plant Science

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